Advanced stage glioma is the most aggressive form of malignant brain tumors with a short survival time. Real-time pathology assisted, or image guided surgical procedures that eliminate tumors promise to improve the clinical outcome and prolong the lives of patients. Our work is focused on the development of a rapid and sensitive assay for intraoperative diagnostics of glioma and identification of optical markers essential for differentiation between tumors and healthy brain tissues. We utilized fluorescence lifetime imaging (FLIM) of endogenous fluorophores related to metabolism of the glioma from freshly excised brains tissues. Macroscopic time-resolved fluorescence images of three intracranial animal glioma models and surgical samples of patients’ glioblastoma together with the white matter have been collected. Several established and new algorithms were applied to identify the imaging markers of the tumors. We found that fluorescence lifetime parameters characteristic of the glioma provided background for differentiation between the tumors and intact brain tissues. All three rat tumor models demonstrated substantial differences between the malignant and normal tissue. Similarly, tumors from patients demonstrated statistically significant differences from the peritumoral white matter without infiltration. While the data and the analysis presented in this paper are preliminary and further investigation with a larger number of samples is required, the proposed approach based on the macroscopic FLIM has a high potential for diagnostics of glioma and evaluation of the surgical margins of gliomas.

The aim of the study was to evaluate the performance of trans-serosal multimodal OCT (MM OCT) in in vivo detecting of changes in microstructure and blood circulation of the small intestine wall caused by arteriovenous ischemia resulted from intestine strangulation. Materials and Methods. In experiments on Wistar rats (n=22), we examined the small intestine wall in vivo using MM OCT; the access to the intestine was reached through laparotomy. The microvasculature and microstructure of the wall were studied before and after acute arteriovenous ischemia created by ligation of a small bowel segment. The results were then added with data obtained from histological and intravital microscopic examination. Results. Trans-serous MM OCT allowed us to visualize the bowel wall to its entire thickness, distinguish between the serous-muscular and mucous-submucosal layers, and detect the villi and functioning blood vessels. The structures were best seen after a fat emulsion had been administered into the bowel lumen. In OCT images made in the optical coherent angiography (OCA) mode, large paired vessels (arteries and veins) and micro-vessels with a diameter of >15 μm could be seen. Most of the blood vessels were imaged in the depth range of 80-300 μm from the surface. Capillaries with a diameter of 7-10 μm were not seen, but they produced an overall bright background. In the OCA images reconstructed from a volume of 2.4×2.4×1.8 mm, the total length of the vascular bed before ischemia was 18.3 [16.6; 19.8] mm. Strangulation of the intestinal loop was associated with changes in the CP OCT picture: the villi-associated vertical pattern and shadows of blood vessels disappeared and the depth of tissue visualization in the cross-channel decreased. The optical equivalents of the serous-muscular layer were preserved; after 180±12 min of ischemia, their proportion in the intestinal wall thickness increased from 25 [18; 32] to 42 [31; 55]% (p=0.031). At that time-point, OCA images of the strangulated bowel loop looked all similar: a uniform dark background with isolated fragmentary large vessels and no signs of blood flow in the microvascular network. Conclusion. Trans-serous MM OCT provides for in vivo visualization of microstructures critical for surgical gastroenterology: the intestinal wall layers including villi and blood vessels of each layer, as confirmed by histological analysis. Destructive processes in the intestinal wall resulting from bowel ligation bring about optical changes, which can be detected using real-time MM OCT.

Introduction: Despite the introduction of increasingly multifaceted diagnostic techniques and the general advances in emergency abdominal and vascular surgery, the outcome of treatment of patients with acute impaired intestinal circulation remains unsatisfactory. The non-invasive and high-resolution technique of optical coherence tomography (OCT) can be used intraoperatively to assess intestine viability and associated conditions that frequently emerge under conditions of impaired blood circulation. This study aims to demonstrate the effectiveness of multimodal (MM) OCT for intraoperative diagnostics of both the microstructure (cross—polarization OCT mode) and microcirculation (OCT angiography mode) of the small intestine wall in patients with acute mesenteric ischemia (AMI). Methods and Participants: A total of 18 patients were enrolled in the study. Nine of them suffered from AMI in segments II-III of the superior mesenteric artery (AMI group), whereby the ischemic segments of the intestine were examined. Nine others were operated on for adenocarcinoma of the colon (control group), thus allowing areas of their normal small intestine to be examined for comparison. Data on the microstructure and microcirculation in the walls of the small intestine were obtained intraoperatively from the side of the serous membrane using the MM OCT system (IAP RAS, Russia) before bowel resection. The MM OCT data were compared with the results of histological examination. Results: The study finds that MM OCT visualized the damage to serosa, muscularis externa, and blood vessels localized in these layers in 100% of AMI cases. It also visualized the submucosa in 33.3% of AMI cases. The MM OCT images of non-ischemic (control group), viable ischemic, and necrotic small intestines (AMI group) differed significantly across stratification of the distinguishable layers, the severity of intermuscular fluid accumulations, and the type and density of the vasculature. Conclusion: The MM OCT diagnostic procedure optimally meets the requirements of emergency surgery. Data on the microstructure and microcirculation of the intestinal wall can be obtained simultaneously in real time without requiring contrast agent injections. The depth of visualization of the intestinal wall from the side of the serous membrane is sufficient to assess the volume of the affected tissues. However, the methodology for obtaining MM OCT data needs to be improved to minimize the motion artefacts generated in actual clinical conditions.

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Development of the novel diagnostic and therapeutic approaches in neuro-oncology requires tumor models that closely reproduce the biological features of patients’ tumors. Patient-derived xenografts (PDXs) are recognized as a valuable and the most “close-to-patient” tool for preclinical studies. However, their establishment is complicated by the factors related to both the surgical material and technique of the orthotopic implantation. The aim of this work was to develop a patient-derived glioblastoma multiform (GBM) model that stably co-expresses luciferase and a far-red fluorescent protein for monitoring of tumor progression in the brain and, using this model, to validate new diagnostic methods—macroscopic fluorescence lifetime imaging (macro-FLIM) and cross-polarization optical coherence tomography (CP OCT). The established model was similar to the original patient’s GBM in terms of histological and immunohistochemical features and possessed reproducible growth in nude mice, which could be observed by both fluorescence and bioluminescence imaging. Our results demonstrated the high potential of macro-FLIM and CP OCT for intraoperative differentiation of GBM from the white matter. Thus, the dual-labeled PDX model of GBM proved to be an excellent approach for observation of tumor development by optical methods.